CHROMATIN STRUCTURE AND TRANS-ACTIVATION OF HUMAN GLOBIN GENES

染色质结构和反式激活是基因表达的两个重要方面 调控 来自活跃转录基因的染色质呈现出一种结构 与非转录基因不同。 已经显示 反式作用蛋白质可以与DNA结合并赋予独特的染色质 这种蛋白质与DNA的相互作用被认为是 基因调控的重要作用。 为了调查 人类珠蛋白基因表达调控机制的两种途径 1）我们研究了细胞核周围的染色质结构， 用DNA酶I和S1核酸酶在K562细胞中扩增人β-珠蛋白基因。 尽管β-珠蛋白基因不表达，我们发现核酸酶 在其5'和3'侧翼区都存在超敏位点。 而3' 超敏位点与活性β珠蛋白基因的超敏位点相同， 5'超敏感位点进一步向上游移动。 这表明 K562细胞中β-珠蛋白表达的缺乏可能与独特的 染色质结构的5'侧翼区域的一个或多个方面， β-珠蛋白基因 2)为了研究反式激活的机制， 在瞬时表达系统中测试珠蛋白启动子。 我们发现 猴珠蛋白启动子可被SV 40 T抗原激活 肾CV-1细胞。 这种反式激活可以进一步刺激， 在珠蛋白质粒中包括SV 40增强子。 的机理 反式激活目前正在研究中。

Chromatin structure and trans-activation are two important aspects of gene regulation. Chromatin from actively transcribed genes exhibits a structure different from that of non-transcribed genes. It has been shown that trans-acting proteins can bind to DNA and confer a unique chromatin structure, and such a protein DNA interaction has been suggested to play an important role in gene regulation. In an attempt to investigate the mechanisms of regulation of human globin gene expression two approaches have been used: 1) We have studied the chromatin structure around the human beta-globin gene in K562 cells by using DNase I and S1 nuclease. Despite the beta-globin gene not being expressed, we found nuclease hypersensitive sites in both its 5' and 3' flanking regions. While the 3' hypersensitive sites are identical to that of active beta globin genes, the 5' hypersensitive sites shift further upstream. This suggests that the lack of beta-globin expression in K562 cells may be related to unique aspect(s) of the chromatin structure in the immediate 5' flanking region of the beta-globin gene. 2) To investigate the mechanism of trans-activation, globin promoters were tested in a transient expression system. We found that the globin promoter can be activated by SV40 T antigen in monkey kidney CV-1 cells. This trans-activation can be further stimulated by including a SV40 enhancer in the globin plasmid. The mechanism of the trans-activation is currently under study.